The long-term goal of our research is to elucidate molecular mechanisms mediating host-microbe interactions and to learn how microbial processes affect the outcome of these relationships. The model bacterium we use for these studies is Xenorhabdus nematophila, which is both a mutualist (of nematodes) and a pathogen (of insects), and therefore an excellent model to understand both types of relationships. This proposal focuses on the mutualism between X. nematophila and the nematode and complements separately funded work in our lab on X. nematophila pathogenesis. Examining the mutualistic interactions of X. nematophila and its nematode host will expand our understanding of how animals evolve and maintain specific and dependent relationships with their micro-flora. The specific aims of this proposal are geared toward genetically and biochemically characterizing several of the colonization initiation factors we have identified, as well as their interactions with each other. Specific Aim 1 will test our hypotheses that membrane-localized colonization factors function in adherence to nematode cells, metal transport, and/or signal transduction. The results of these studies will elucidate the roles of Nil (nematode intestine localization) proteins in colonization events, shed light on the possible functions of Nil homologs of gram-negative pathogens in initiating infections, and further our understanding of molecular mechanisms mediating host-range specificity. Specific Aim 2 is designed to dissect the functions of two regulators required for colonization, a sigma factor and a regulatory RNA. These experiments will yield insight into the signals perceived by X. nematophila during colonization, further define a colonization-specific regulon, and expand our knowledge of the role of a ubiquitous transcription factor in the natural ecology of a bacterium. [unreadable] [unreadable]